MMP-9 is an endopeptidase playing important role in neuronal plasticity. Although multiple factors regulating MMP-9 expression have been described in different cell types, the molecular mechanism directly controlling its transcription in neurons remains poorly understood. The aim of the current study was to determine, whether SRF/c-Fos pathway is involved in the transcriptional regulation of MMP-9 in neurons. Real-Time PCR analysis revealed strong upregulation of MMP-9 mRNA levels after stimulation of rat primary cortical neurons with BDNF. Additionally, elevated MMP-9 gelatinolytic activity was observed. To investigate mechanism of MMP-9 promoter regulation, we used luciferase gene reporter assay system in which luciferase gene is controlled by MMP-9 promoter fragment (-1369/+35). Treatment of neurons with BDNF led to MMP-9 promoter activation, that was dependent on ERK1/2 actvity, as demonstrated using selective inhibitor or overexpressing constitutively active MKK1. As in MMP-9 promoter there are two AP-1 binding sites, we investigated whether AP-1 contributes to the BDNF-mediated MMP-9 transcription in neurons. MMP-9 reporter construct was induced upon overexpression of different AP-1 dimers in neurons, the most potent being those containing c-Fos. Moreover, BDNFinduced activation of the MMP-9 reporter construct was reduced if proximal, but not distal, AP-1 binding site was mutated. Furthermore knocking-down c-Fos expression in neurons by shRNA decreased MMP-9 gene activation in response to BDNF. As c-fos gene is a known target of SRF, we tested whether SRF can contribute to MMP-9 transcription. Inhibition of SRF by the overexpression of dominant-negative mutant of SRF or using shRNA targeting SRF, abolished BDNF-induced activation of MMP-9 promoter. Our data indicate that MMP-9 expression in neurons can be induced by BDNF. The signal propagation could involve ERK1/2 pathway and SRF-mediated transcription of c-fos gene resulting in activation of MMP-9 promoter.
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